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BACKGROUND AND AIM Previous two generations of esophageal capsule did not show adequate detection rates for Barrett's esophagus (BE). We assessed the diagnostic accuracy of a novel third generation capsule with an improved frame rate of 35 frames per second for the detection of BE in a pilot study. METHODS This was a blinded prospective pilot study conducted at a tertiary medical center. Patients with known BE (at least C0M > 1) who presented for endoscopic surveillance (May to October 2017) were included. All patients underwent novel esophageal capsule (PillCam™ UGI; Medtronic) ingestion using the simplified ingestion protocol followed by standard high-definition upper endoscopy (esophagogastroduodenoscopy [EGD]). Capsule endoscopy findings were interpreted by examiners blinded to endoscopy results and compared with endoscopic findings (gold standard). Following completion of both tests, a subjective questionnaire was provided to all patients regarding their experience. RESULTS Twenty patients (95%males, mean age 66.3 [±7.9] years) with BE undergoing surveillance EGD were eligible. The mean BE length was 3.5 (±2.7) cm. Novel esophageal capsule detected BE in 75% patients when images were compared with endoscopy. Novel capsule detected BE in 82% patients when the BE length was ≥2 cm. The mean esophageal transit time was 0.59 s. On a subjective questionnaire, all 20 patients reported novel capsule as being more convenient compared with EGD. CONCLUSIONS In this pilot, single-center study, novel esophageal capsule was shown to be not ready for population screening of BE. Studies integrating artificial intelligence into improved quality novel esophageal capsule should be performed for BE screening.

BACKGROUND & AIMS: European guidelines recommend different surveillance intervals of non-dysplastic Barrett’s esophagus (NDBE) based on segment length, as opposed to guidelines in the United States, which do recommend surveillance intervals based on BE length. We studied rates of progression of NDBE to high-grade dysplasia (HGD) or esophageal adenocarcinoma (EAC) in patients with short-segment BE using the definition of BE in the latest guidelines (length ≥1 cm). METHODS: We collected demographic, clinical, endoscopy, and histopathology data from 1883 patients with endoscopic evidence of NDBE (mean age, 57.3 years; 83.5% male; 88.1% Caucasians) seen at 7 tertiary referral centers. Patients were followed for a median 6.4 years. Cases of dysplasia or EAC detected within 1 year of index endoscopy were considered prevalent and were excluded. Unadjusted rates of progression to HGD or EAC were compared between patients with short (≥1 and

Summary Background and Aims: As obtaining adequate tissue on biopsy is critical for the detection of residual and recurrent intestinal metaplasia/dysplasia in Barrett’s esophagus (BE) patients undergone Barrett’s endoscopic eradication therapy (BET), we decided to compare the adequacy of biopsy specimens using jumbo versus standard biopsy forceps. Methods: This is a two-center study of patients’ post-radiofrequency ablation of dysplastic BE. After BET, jumbo (Boston Scientific©, Radial Jaw 4, opening diameter 2.8 mm) or standard (Boston Scientific©, Radial Jaw 4, opening diameter 2.2 mm) biopsy forceps were utilized to obtain surveillance biopsies from the neo-squamous epithelium. Presence of lamina propria and proportion of squamous epithelium with partial or full thickness lamina propria was recorded by two experienced gastrointestinal pathologists who were blinded. Squamous epithelial biopsies that contained at least two-thirds of lamina propria were considered ‘adequate’. Results: In a total of 211 biopsies from 55 BE patients, 145 biopsies (29 patients, 18 males, mean age 61 years, interquartile range [IQR] 33–83) were obtained using jumbo forceps and 66 biopsies (26 patients, all males, mean age 65 years, IQR 56–76) using standard forceps biopsies. Comparing jumbo versus standard forceps, the proportion of specimens with any subepithelial lamina propria was 51.7% versus 53%, P = 0.860 and the presence of adequate subepithelial lamina propria was 17.9% versus 9.1%, P = 0.096 respectively. Conclusions: Use of jumbo forceps does not appear to have added advantage over standard forceps to obtain adequate biopsy specimens from the neo-squamous mucosa post-ablation.

AIM To evaluate annual incidence of low grade dysplasia (LGD) progression to high grade dysplasia (HGD) and/or esophageal adenocarcinoma (EAC) when diagnosis was made by two or more expert pathologists. METHODS Studies evaluating the progression of LGD to HGD or EAC were included. The diagnosis of LGD must be made by consensus of two or more expert gastrointestinal pathologists. Articles were searched in Medline, Pubmed, and Embase. Pooled proportions were calculated using fixed and random effects model. Heterogeneity among studies was assessed using the I2 statistic. RESULTS Initial search identified 721 reference articles, of which 53 were selected and reviewed. Twelve studies (n = 971) that met the inclusion criteria were included in this analysis. Among the total original LGD diagnoses in the included studies, only 37.49% reached the consensus LGD diagnosis after review by two or more expert pathologists. Total follow up period was 1532 patient-years. In the pooled consensus LGD patients, the annual incidence rate (AIR) of progression to HGD and or EAC was 10.35% (95%CI: 7.56-13.13) and progression to EAC was 5.18% (95%CI: 3.43-6.92). Among the patients down staged from original LGD diagnosis to No-dysplasia Barrett’s esophagus, the AIR of progression to HGD and EAC was 0.65% (95%CI: 0.49-0.80). Among the patients down staged to Indefinite for dysplasia, the AIR of progression to HGD and EAC was 1.42% (95%CI: 1.19-1.65). In patients with consensus HGD diagnosis, the AIR of progression to EAC was 28.63% (95%CI: 13.98-43.27). CONCLUSION When LGD is diagnosed by consensus agreement of two or more expert pathologists, its progression towards malignancy seems to be at least three times the current estimates, however it could be up to 20 times the current estimates. Biopsies of all Barrett’s esophagus patients with LGD should be reviewed by two expert gastroenterology pathologists. Follow-up strict surveillance programs should be in place for these patients.

Identification of the human microbiome has proven to be of utmost importance with the emerging role of bacteria in various physiological and pathological processes. High throughput sequencing strategies have evolved to assess the composition of the microbiome. To identify possible bias that may exist in the processing of tissue for whole genome sequencing (WGS), it is important to evaluate the extraction method on the overall microbial content and composition. Here we compare two different methods of extraction, homogenization vs. enzymatic lysis, on gastric, esophageal and colorectal biopsies and survey the microbial content and composition using WGS and quantitative PCR (qPCR). We examined total bacterial content using universal 16S rDNA qPCR as well as the abundance of three phyla (Actinobacter, Firmicutes, Bacteroidetes) and one genus (Fusobacterium). We found minimal differences between the two extraction methods in the overall community structure. Furthermore, based on our qPCR analysis, neither method demonstrated preferential extraction of any particular clade of bacteria, nor significantly altered the detection of Gram-positive or Gram-negative organisms. However, although the overall microbial composition remained very similar and the most prevalent bacteria could be detected effectively using either method, the precise community structure and microbial abundances between the two methods were different, primarily due to variations in detection of low abundance genus. We also demonstrate that the homogenization extraction method provides higher microbial DNA content and higher read counts from human tissue biopsy samples of the gastrointestinal tract.

BACKGROUND Validated probe-based confocal endomicroscopy (pCLE) criteria for distinguishing hyperplastic polyps (HPs) and tubular adenomas (TA) have not yet been developed. AIM To develop pCLE criteria for distinguishing HP from TA and evaluate its performance characteristics among experts. METHODS pCLE criteria for colon polyp histology were developed and tested in 2 phases prospectively. Phase I: 8 preliminary criteria were developed and tested internally. Criteria achieving an accuracy of >75% (epithelial surface: regular vs. irregular; goblet cells: increased vs. decreased; gland axis: horizontal vs. vertical; gland shape: slit/stellate vs. villiform; image scale: gray vs. dark) were evaluated in Phase II of study wherein external assessors evaluated these criteria in a separate set of pCLE videos. Accuracy and interobserver agreement (95% confidence intervals) were determined for colon histology prediction. RESULTS Phase I (criteria development/internal testing): 8 criteria were assessed by 4 pCLE experts using 28 videos (14 HP/14 TA). Five of 8 pCLE criteria met selection for phase II (accuracy >75%). Phase II (external validation): 36 pCLE colon polyp videos (HP 16/TA 20) were evaluated by 8 external assessors. Overall accuracy in diagnosis of colon polyp histology was 84.9% (95% confidence interval, 81.7-87.7). Of predictions made with high confidence (75%), histology was predicted with an accuracy of 91%, sensitivity 83%, specificity 100%, negative predictive value 87% and positive predictive value 98%. Interobserver agreement was substantial (κ=0.73). CONCLUSIONS We demonstrate the development and validation of pCLE criteria for prediction of colon polyp histology. Using these criteria, overall accuracy in differentiating TA from HP was high with substantial interobserver agreement.

Many patients with a1 cm segment of columnar metaplasia in the distal esophagus, also called an irregular Z line, are encountered. These patients, often referred to as patients with Barrett's esophagus (BE), are enrolled in surveillance programs. However, little is known about their risk of high-grade dysplasia (HGD) or esophageal adenocarcinoma (EAC). We aimed to determine the incidence of HGD and EAC in patients with irregular Z line with intestinal metaplasia.We performed a prospective, multicenter cohort study of patients who underwent endoscopic examination for BE at tertiary care referral centers in the United States and Europe. We analyzed data from 1791 patients (mean age, 56 ± 17 years) found to have non-dysplastic BE at the index endoscopy and after 1 year or more of follow-up. Patients were followed for a median of 5.9 years (interquartile range, 3.1-8.3 years). We calculated rates of progression to HGD or EAC between groups of patients with irregular Z line (n = 167) and those with BE of ≥ 1 cm (n = 1624).A higher proportion of patients in the irregular Z-line group were female (26.3%) than in the BE group (14.8% female BE) (P.001). A lower proportion of patients in the irregular Z-line group were smokers (33.5%) than in the BE group (52.6% smokers). None of the patients with irregular Z line developed HGD or EAC during a median follow-up period of 4.8 years (interquartile range, 3.2-8.3 years). All 71 incident cases of HGD or EAC developed in patients with BE of ≥1 cm in length. On multivariate analysis, patients with irregular Z line and patients with BE of ≥ 1 cm did not differ significantly in age, race, or duration of follow-up.In a prospective, multicenter cohort study, we found that patients with irregular Z line do not develop HGD or esophageal cancer within 5 years after index endoscopy.

Although Barrett’s esophagus is the primary precursor of esophageal adenocarcinoma, the annual incidence of adenocarcinoma is 0.12% in a Barrett’s esophagus population. We hypothesized that the esophageal microbiome may be associated with mucosal immunity changes that may determine subsequent carcinogenesis. To better understand the association among the local microbiota composition, immune response and progression of esophageal cancer, we performed whole genome sequencing (WGS) and bulk RNASeq in parallel, from endoscopic biopsies (n=23) collected from patients with normal squamous cell (n=9), non-dysplastic Barrett’s esophagus (n=8) and esophageal adenocarcinoma (n = 6). Using a novel customized computational pipeline to identify and characterize bacteria from low microbial content endoscopic samples (Zhang et al, Genome Biology, 2015). WGS analysis revealed a higher microbial diversity in normal squamous cell carcinomas compared to the non-dysplastic Barrett’s esophagus with high abundance of Veilonella Parvula, Prevotella melaninogenica and Streptococcus parasanguinis. Unsupervised clustering analysis of the entire data set revealed high abundance of Fusobacterium nucleatum in adenocarcinoma samples, which has been associated with a shorter survival in esophageal cancer. To characterize the immune infiltration in the mucosal biopsy samples, we defined a 784-gene immune panel. Unsupervised clustering analysis of gene expression revealed a much higher immune infiltration in most adenocarcinoma and some non-dysplastic Barrett’s compared with normal esophageal squamous tissue samples. Especially CD4+ Th1 and Th2 helper cell as well as CD8+ T cell associated immune response are highly enriched in some adenocarcinoma and non-dysplastic Barrett’s esophageal samples. To further investigate the effect of immune tumor microenvironment on efficacy of immunotherapy, we performed single cell sequencing on matching normal and tumor tissues collected at baseline and post-treatment from 6 patients subjected to anti-PD1 therapy. Using this approach, we can identify both innate and adaptive immune cells in both pre and post treatment biopsies. We specifically observe upregulation of MHCII associated cells in post-treatment biopsies. Together, our data suggest that esophageal cancers display distinct microbial patterns associated with chronic inflammation and a tumor-promoting pro-inflammatory microenvironment. Note: This abstract was not presented at the meeting. Citation Format: Chao Zhang, Prashant V. Thakkar, Prateek Sharma, Sreekar Vennelaganti, Doron Betel, Manish A. Shah. Understanding associations among local microbiome, immune response, and efficacy of immunotherapy in esophageal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2826.

Background and Aims Proximal colon adenomas can be missed during routine colonoscopy. Use of a cap or hood on the tip of the colonoscope has been shown to improve overall adenoma detection with variable rates. However, it has not been systematically evaluated for detection of proximal colon or right-sided adenomas where the cap may have maximum impact on adenoma detection rate (ADR). Our aim was to perform a systematic review and meta-analysis to evaluate the impact of cap-assisted colonoscopy (CC) on right-sided ADRs (r-ADRs) compared with standard colonoscopy (SC). Methods PubMed, EMBASE, SCOPUS, and Cochrane databases as well as secondary sources (bibliographic review of selected articles and major GI proceedings) were searched through October 1, 2016. Primary outcome was the pooled rate of r-ADR. Detection of flat adenoma, sessile serrated adenoma/polyp (SSA/P), and number of right-sided adenomas per patient were also assessed. Pooled odds ratio (OR) and 95% confidence intervals (CIs) were calculated using random-effect models. Results We screened 686 records and analyzed data from 4 studies (CC group, 2546 patients; SC group, 2547 patients) that met criteria for determination of r-ADRs, whereas 6 studies (CC group, 3159 patients; SC group, 3137 patients) were analyzed to estimate right-sided adenomas per patient. r-ADR was significantly higher with CC compared with SC (23% vs 17%; OR, 1.49; 95% CI, 1.08-2.05; I 2 = 79%; P = .01). CC also improved detection rates of flat adenoma (OR, 2.08; 95% CI, 1.35-3.20; P P = .04). The total number of right-sided adenomas (CC: 1428 [60%] vs SC: 1127 [58%]) and number of right-sided adenomas per patient (CC, .71 ± .5, vs SC, .65 ± .62 [mean ± standard deviation]) were numerically higher for CC but were not statistically significant ( P = .43). Approximately 17 CCs would be required to detect an additional patient with right-sided adenoma. Conclusions Use of CC significantly improves the proximal colon ADR. In addition, flat adenoma and serrated colonic lesion detection rates are also significantly higher as compared with SC.

Background and Aims Focal EMR followed by radiofrequency ablation (f-EMR + RFA) and stepwise or complete EMR (s-EMR) are established strategies for eradication of Barrett's esophagus (BE)-related high-grade dysplasia (HGD) and/or esophageal adenocarcinoma (EAC)/intramucosal carcinoma (IMC). The objective of this study was to derive pooled rates of efficacy and safety of individual methods in a large cohort of patients with BE and to indirectly compare the 2 methods. Methods PubMed, Embase, Web of Science, Cochrane, and major conference proceedings were searched. A systematic review and pooled analysis were carried out to determine the following outcomes in patients with BE undergoing either f-EMR + RFA or s-EMR: (1) complete eradication rates of neoplasia (CE-N) and intestinal metaplasia (CE-IM); (2) recurrence rates of cancer (EAC), dysplasia, and IM; (3) incidence rates of adverse events. Mixed logistic regression was performed as an exploratory analysis to examine differences in outcomes between the 2 methods. Results Nine studies (774 patients) of f-EMR + RFA and 11 studies (751 patients) of s-EMR were included. Patients undergoing f-EMR + RFA had high BE eradication rates (CE-N, 93.4%; CE-IM, 73.1%), whereas strictures occurred in 10.2%, bleeding in 1.1%, and perforations in 0.2% of patients. Recurrence of EAC, dysplasia, and IM was 1.4%, 2.6%, and 16.1%, respectively, in this group. Patients undergoing s-EMR also showed high BE eradication rates (CE-N, 94.9%; CE-IM, 79.6%) but a higher rate of adverse events (strictures in 33.5%, bleeding in 7.5%, and perforation in 1.3%). Recurrence of EAC, dysplasia, and IM was 0.7%, 3.3%, and 12.1%, respectively, in the s-EMR group. Mixed logistic regression showed that patients undergoing s-EMR might be more likely to develop esophageal strictures (odds ratio [OR], 4.73; 95% confidence interval [CI], 1.61-13.85; P = .005), perforation (OR, 7.00; 95% CI, 1.56-31.33; P = .01), and bleeding (OR, 6.88; 95% CI, 2.19-21.62; P = 0.001) compared with f-EMR + RFA. Conclusions In patients with HGD/EAC, f-EMR followed by RFA seems to be equally effective as and safer than s-EMR.

Gastric and esophageal cancers remain the second and the sixth leading cause of cancer related deaths respectively. Dysbiosis of the gut microbiome remains a strong risk factor for gastrointestinal cancer, likely related to altered mucosal immunity and a pro-inflammatory immune microenvironment. However, the molecular interactions between host human cells and the microbiome are poorly understood as it pertains to gastric and esophageal cancer. Our main goal is to understand how alteration of regional mucosal microbiome alters the tumor microenvironment, thus creating conditions conducive to tumorigenesis. Understanding these changes at the molecular level will help contribute to our knowledge of the disease pathophysiology. Using a novel customized computational pipeline to identify and characterize bacteria from low microbial content endoscopic samples (Zhang et al., Genome Biol 2015), we used low-pass (10-30x) whole genome sequencing (WGS) to examine the microbiome composition of 15 gastric cancer samples along with their adjacent non-cancer mucosal tissue (n=30). We found a higher overall bacterial content in tumor samples compared to that of their corresponding matching normal tissues. Furthermore, we observed a dysbiosis in samples positive for Helicobacter pylori, consistent with previous findings that H. pylori alters composition of the gastric microbiome. Although a wide variety of other microbes were detected, we found specific enrichment of organisms like Veilonella Parvula (12/15) and Prevotella melaninogenica (10/15) in tumor tissues, both of which have been implicated in tumorigenicity. A subset of these samples (10 gastric cancer and 12 adjacent normal) underwent parallel RNAseq analysis and using a gene expression signature panel of 176 genes compiled from several published series, we examined immune cell types, specifically B cells, CD8+ T cells, Treg cells, CD4+ Th1, Th2 and Th17 helper cells and macrophages, to determine an association with tumor versus the normal mucosa. We found an overall higher immune response, and more specifically a higher association of Th1 and Th2 helper cells as well as macrophages, with tumors as compared to normal mucosa. We further corroborated these findings using a single cell RNA sequencing approach on three tumor and matching normal samples. Using ELISA, we detected a significantly increased expression of pro-inflammatory cytokines such as TNFα, IL-8, GRO, MCP-1 and IL-1a in tumor samples than in normal mucosa. Increased expression of TNF-α and IL-8 has been directly correlated to tumor invasion and metastasis, and tumor vascularity respectively. Additionally, MCP-1 has been shown to be important in recruitment of macrophages and is secreted by gastric epithelial cells in response to pro-inflammatory cytokines upon H. pylori infection. We used a similar approach, of performing WGS and RNAseq analysis in parallel, on 9 normal squamous cell carcinoma, 8 non-dysplastic Barrett’s esophagus and 6 esophageal adenocarcinoma samples (n = 23 samples). WGS analysis revealed a higher microbial diversity in normal squamous cell carcinomas compared to the non-dysplastic Barrett’s esophagus with high abundance of Veilonella Parvula, Prevotella melaninogenica and Streptococcus parasanguinis. Unsupervised clustering analysis of the entire data set revealed high abundance of Fusobacterium nucleatum in adenocarcinoma samples, which has been associated with a shorter survival in Esophageal cancer. Unsupervised clustering analysis of RNAseq results revealed a high enrichment of CD4+ Th1 and Th2 helper cell as well as CD8+ T cell associated immune response in some adenocarcinoma and non-dysplastic Barrett’s esophageal samples. Together, our data are derived from endoscopic biopsy samples from patients, and suggest that both gastric and esophageal cancers display distinct microbial patterns associated with chronic inflammation and a tumor-promoting pro-inflammatory microenvironment. Citation Format: Prashant V. Thakkar, Chao Zhang, Prateek Sharma, Sreekar Vennelaganti, Doron Betel, Manish A. Shah. Understanding the association of gut microbiota and tumor microenvironment in gastric and esophageal cancer [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2017 Oct 1-4; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2018;6(9 Suppl):Abstract nr B39.

Gastric and Esophageal cancers remain one of the leading causes of cancer related deaths. Dysbiosis of gut microbiome is a strong risk factor for gastrointestinal cancer, likely due to altered mucosal immunity and pro-inflammatory immune microenvironment. However, molecular interactions between host human cells and microbiome are poorly understood in context of gastric and esophageal cancer. Our main goal is to understand how altered mucosal microbiome alters tumor microenvironment, thus creating conditions conducive to tumorigenesis. Using a novel computational pipeline combined with low-pass (10-30x) whole genome sequencing (WGS), we examined microbiome composition of 15 gastric cancer (GC) and adjacent normal tissue (n=30). We found a higher overall bacterial content in tumor samples compared to that of matching normal tissues. Furthermore, we observed dysbiosis in H. pylori positive samples, consistent with previous findings. Among other microbes, we found specific enrichment of V. Parvula (12/15) and P. melaninogenica (10/15) in tumor tissues, both of which have been implicated in tumorigenicity. A subset of these samples (10 GC and 12 normal) underwent parallel RNAseq analysis. Using gene expression signature panel of 176 genes, we examined association of various immune cell types with tumor versus normal mucosa. We found an overall higher immune response, and higher association of Th1 and Th2 helper cells and macrophages, with tumors as compared to normal mucosa. We further corroborated these findings using single cell RNAseq approach. Using ELISA, we detected significantly increased expression of pro-inflammatory cytokines such as TNF-α, IL-8, GRO, MCP-1 and IL-1a in tumor samples than normal mucosa. Increased TNF-α and IL-8 expression has been directly correlated to tumor invasion and metastasis, and tumor vascularity respectively. Additionally, MCP-1 is known to recruit macrophages and is secreted by gastric epithelial cells in response to pro-inflammatory cytokines upon H. pylori infection. Using a similar approach, we performed WGS and RNAseq, on 9 normal squamous cell carcinoma (NSCC), 8 non-dysplastic Barrett's esophagus (BE) and 6 esophageal adenocarcinoma samples. WGS analysis revealed higher microbial diversity in NSCCs compared to BE samples with high abundance of V. Parvula, P. melaninogenica and S. parasanguinis. Unsupervised clustering analysis revealed high abundance of F. nucleatum in adenocarcinoma, which has been associated with shorter survival in esophageal cancer. Unsupervised clustering analysis of RNAseq data revealed high enrichment of CD4+ Th1 and Th2 helper cell and CD8+ T cell associated immune response in some adenocarcinoma and BE samples. Together our data suggest that both gastric and esophageal cancers display distinct microbial patterns associated with chronic inflammation and tumor-promoting pro-inflammatory microenvironment. Citation Format: Prashant V. Thakkar, Chao Zhang, Prateek Sharma, Sreekar Vennelaganti, Doron Betel, Manish A. Shah. Understanding the association of gut microbiota and tumor microenvironment in gastric and esophageal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5128.